The objective is to develop a molecular understanding of sodium-coupled amino acid transport by the small intestinal epithelium. Ion coupled solute transport plays an essential role in gastrointestinal function, yet basic knowledge of transport carriers is incomplete or lacking. This study is designed to address this problem by focusing on the IMINO cotransporter, a brush border membrane carrier with an exceptionally selective substrate requirement for L-proline and Na+. This ideal prototype cotransport carrier is readily studied in brush border membrane vesicles isolated from rabbit jejunal enterocytes. Experimentally, sodium-coupled proline uptake into brush border vesicles will be measured using radioactive tracers, and potential-sensitive fluorescent dye optical techniques. Transport will be investigated with respect to (1) the molecular mechanism of transport events, and (2) the carrier's structure/function relationship as probed by high-energy electron inactivation: (1) Transport kinetics of initial influx will be measured under various conditions of cis and trans substrates, and with imposed membrane PD. The kinetics will be evaluated in light of known mechanistic paradigms, and will provide a comprehensive mathematical model which describes sodium-coupled proline transport in the terms of an enzyme-like reaction mechanism. (2) The membrane-bound in situ molecular weight of the functional proline carrier and its possible subunits will be probed utilizing high-energy electron radiation inactivation, with application of target theory. The project holds long-term health-related implications. It serves as a prototype, so that the mechanisms of the other known amino acid carriers can be investigated in both brush border and basolateral membrane vesicles. This will permit a composite molecular description of amino acid transcellular absorption in healthy intestine, will provide a foundation for understanding ion-coupled solute transport, and thus will enhance the understanding of the membrane pathophysiology of genetic defects of amino acid transport in intestinal and renal malabsorption syndromes.